At least two independent genetic systems, nuclear and mitochondrial, contribute to the biogenesis of mitochondria in yeast and to the maintenance of its semiautonomous existence. Conditional, temperature sensitive (ts) and cold sensitive (cs) nuclear and mitochondrial mutants affecting mitochondrial respiration and function will be isolated and characterized genetically and biochemically. The newly isolated ts and cs mitochondrially inherited mutants will be mapped relative to the established antibiotic resistance markers. The coding capacity of the mitochondrial genome will also be determined in vitro by the coupled transcription-translation of purified mitochondrial DNA (mDNA) and by the translation of mitochondrial specific messenger RNA. At the biochemical level, a study will be made of the combined expression of the nuclear and mitochondrial genetic systems that contribute to the structure and organization of the enzyme complexes of the inner mitochondrial membrane. Second site nuclear (and possibly mitochondrial) suppressor mutations of mitochondrially inherited ts and cs lesions should provide an insight into the contribution of nuclear and mitochondrial gene products to the mitochondrial protein synthesizing machinery. In addition, nuclear conditional mutants will be isolated that affect specifically mitochondrial nucleic acid synthesis. The method is based on the selective labelling of mDNA with bromouracil so that subsequent irradiation of a mutagenized culture would select mutants defective in mDNA synthesis. The nuclear and mitochondrial mutants should prove valuable in the role that each system plays in mitochondrial biogenesis.